1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-diones and their use for the treatment of pain

ABSTRACT

Compounds according to structural diagram (I) 
                 
         are disclosed, wherein R 1 , A, E and D are as defined in the specification. Also disclosed are pharmaceutical compositions comprising a pain-ameliorating effective amount of a compound in accord with structural diagram (I).       

     The following is an examiner&#39;s statement of reasons for allowance: In view of applicants&#39; amendments to the claims which limit “E” to phenyl or cycloalkyl groups the rejections of the previous action no longer pertain as they at best teach “E” as alkyl.

This is the National Stage of International Application PCT/SE01/02126,filed Sep. 28, 2001, which claims priority under 35 U.S.C. §119(e) ofU.S. Provisional Applications 60/171,906, filed Dec. 23, 1999 and60/236,880, filed Sep. 29, 2000.

FIELD OF THE INVENTION

This invention relates to the treatment or prevention of pain ornociception.

RELATED ART

Pain is a sensory experience distinct from sensations of touch,pressure, heat and cold. It is often described by sufferers by suchterms as bright, dull, aching, pricking, cutting or burning and isgenerally considered to include both the original sensation and thereaction to that sensation. This range of sensations, as well as thevariation in perception of pain by different individuals, renders aprecise definition of pain difficult, however, many individuals sufferwith severe and continuous pain.

Pain that is caused by damage to neural structures is often manifest asa neural supersensitivity or hyperalgesia and is termed “neuropathic”pain. Pain can also be “caused” by the stimulation of nociceptivereceptors and transmitted over intact neural pathways, such pain istermed “nociceptive” pain.

The level of stimulation at which pain becomes noted is referred to asthe “pain threshold.” Analgesics are pharmaceutical agents which relievepain by raising the pain threshold without a loss of consciousness.After administration of an analgesic drug a stimulus of greaterintensity or longer duration is required before pain is experienced. Inan individual suffering from hyperalgesia an analgesic drug may have ananti-hyperalgesic effect. In contrast to analgesics, agents such aslocal anaesthetics block transmission in peripheral nerve fibers therebyblocking awareness of pain. General anaesthetics, on the other hand,reduce the awareness of pain by producing a loss of consciousness.

Tachykinin antagonists have been reported to induce antinociception inanimals, which is believed to be analogous to analgesia in man (Maggi etal, J. Auton. Pharmacol. (1993) 13, 23-93). In particular, non-peptideNK-1 receptor antagonists have been shown to produce such analgesia. Forexample, the NK-1 receptor antagonist RP 67,580 produced analgesia withpotency comparable to that of morphine (Garret et al, Proc. Natl. Acad.Sci. USA (1993) 88, 10208-10212).

The opioid analgesics are a well-established class of analgesic agentswith morphine-like actions. Synthetic and semi-synthetic opioidanalgesics are derivatives of five chemical classes of compound:phenanthrenes; phenylheptylamines; phenylpiperidines; morphinans; andbenzomorphans. Pharmacologically these compounds have diverseactivities, thus some are strong agonists at the opioid receptors (e.g.morphine); others are moderate to mild agonists (e.g. codeine); stillothers exhibit mixed agonist-antagonist activity (e.g. nalbuphine); andyet others are partial agonists (e.g. nalorphine). Whilst an opioidpartial agonist such as nalorphine, (the N-alkyl analogue of morphine)will antagonize the analgesic effects of morphine, when given alone itcan be a potent analgesic in its own right.

Of all of the opioid analgesics, morphine remains the most widely used,but, in addition to its therapeutic properties, it has a number ofdrawbacks including respiratory depression, decreased gastrointestinalmotility (resulting in constipation), nausea and vomiting. Tolerance andphysical dependence also limit the clinical uses of opioid compounds.

Aspirin and other salicylate compounds are frequently used in treatmentto interrupt amplification of the inflammatory process in rheumatoiddiseases and arthritis and temporarily relieve the pain. Other drugcompounds used for these purposes include phenylpropionic acidderivatives such as Ibuprofen and Naproxen, Sulindac, phenyl butazone,corticosteroids, antimalarials such as chloroquine andhydroxychloroquine sulfate, and fenemates (J. Hosp. Pharm., 36:622 (May1979)). These compounds, however, are ineffective for neuropathic pain.

Available therapies for pain also have drawbacks. Some therapeuticagents require prolonged use before an effect is experienced by thepatient. Other existing drugs have serious side effects in certainpatients, and subjects must be carefully monitored to ensure that anyside effects are not unduly threatening. Most existing drugs provideonly temporary relief from pain and must be taken consistently on adaily or weekly basis. With disease progression the amount of medicationneeded to alleviate the pain often increases, thus increasing thepotential for adverse side effects.

NMDA receptors are defined by the binding of N-methyl-D-aspartate (NMDA)comprise a receptor/ion channel complex with several differentidentified binding domains. NMDA itself is a molecule structurallysimilar to glutamate (Glu) which binds at the glutamate binding suiteand is highly selective and potent in activating the NMDA receptor(Watkins (1987); Olney (1989)).

Many compounds are known that bind at the NMDA/Glu binding site (forexample CPP, DCPP-ene, CGP 40116, CGP 37849, CGS 19755, NPC 12626, NPC17742, D-AP5, D-AP7, CGP 39551, CGP-43487, MDL-100,452, LY-274614,LY-233536, and LY233053). Other compounds, referred to asnon-competitive NMDA antagonists, bind at other sites in the NMDAreceptor complex (examples are phencyclidine, dizocilpine, ketamine,tiletamine, CNS 1102, dextromethorphan, memantine, kynurenic acid, CNQX,DNQX, 6,7-DCQX, 6,7-DCHQC, R(+)-HA-966, 7-chloro-kynurenic acid,5,7-DCKA, 5-iodo-7-chloro-kynurenic acid, MDL-28,469, MDL-100,748,MDL-29,951, L-689,560, L-687,414, ACPC, ACPCM, ACPCE, arcaine,diethylenetriamine, 1,10-diaminodecane, 1,12-diaminododecane,ifenprodil, and SL-82.0715). These compounds have been extensivelyreviewed by Rogawski (1992) and Massieu et. al., (1993), and articlescited therein.

In addition to its physiological function, glutamate (Glu) can beneurotoxic. Glu neurotoxicity is referred to as “excitotoxicity” becausethe neurotoxic action of Glu, like its beneficial actions, is mediatedby an excitatory process (Olney (1990); Choi (1992)). Normally, when Gluis released at a synaptic receptor, it binds only transiently and isthen rapidly removed from the receptor by a process that transports itback into the cell. Under certain abnormal conditions, including stroke,epilepsy and CNS trauma, Glu uptake fails and Glu accumulates at thereceptor resulting in a persistent excitation of electrochemicalactivity that leads to the death of neurons that have Glu receptors.Many neurons in the CNS have Glu receptors, so excitotoxicity can causean enormous amount of CNS damage.

Acute excitotoxicity injury can occur as a result of ischemic events,hypoxic events, trauma to the brain or spinal cord, certain types offood poisoning which involve an excitotoxic poison such as domoic acid,and seizure-mediated neuronal degeneration, which can result frompersistent epileptic seizure activity (status epilepticus). A large bodyof evidence has implicated the NMDA receptor as one receptor subtypethrough which Glu mediates a substantial amount of CNS injury, and it iswell established that NMDA antagonists are effective in protecting CNSneurons against excitotoxic degeneration in these acute CNS injurysyndromes (Choi (1988); Olney (1990)).

In addition to neuronal damage caused by acute insults, excessiveactivation of Glu receptors may also contribute to more gradualneurodegenerative processes leading to cell death in various chronicneurodegenerative diseases, including Alzheimer's disease, amyotrophiclateral sclerosis, AIDS dementia, Parkinson's disease and Huntington'sdisease (Olney (1990)). It is generally considered that NMDA antagonistsmay prove useful in the therapeutic management of such chronic diseases.

In the 1980's it was discovered that PCP (also known as “angel dust”)acts at a “PCP recognition site” within the ion channel of the NMDA Glureceptor. PCP acts as a non-competitive antagonist that blocks the flowof ions through the NMDA ion channel. More recently it has becomeevident that drugs which act at the PCP site as non-competitive NMDAantagonists are likely to have psychotomimetic side effects. Further, itis now recognized that certain competitive and non-competitive NMDAantagonists can cause similar pathomorphological effects in rat brain(Olney et. al., (1991); Hargreaves et. al., (1993)). Such compounds alsohave psychotomimetic effects in humans (Kristensen et. al., (1992);Herrling (1994); Grotta (1994)).

The glycine binding site of the NMDA receptor complex is distinguishablefrom the Glu and PCP binding sites. Also, it has recently beendiscovered that NMDA receptors occur as several subtypes which arecharacterized by differential properties of the glycine binding site ofthe receptor. Many compounds that bind at the NMDA receptor glycinesite, useful for the treatment of stroke and neurodegenerativeconditions, have been described in U.S. Pat. Nos. 5,604,227; 5,733,910;5,599,814; 5,593,133; 5,744,471; 5,837,705 and 6,103,721.

SUMMARY OF THE INVENTION

It has now been discovered that certain compounds which exhibit theproperty of binding to the NMDA receptor glycine site have utility forthe amelioration of pain and particularly for the amelioration ofneuropathic pain.

In a first aspect the invention provides compounds according tostructural diagram I useful for the treatment of pain,

wherein: R¹ is halo; A is CH; E is selected from C₁₋₄alkyl, phenyl andC₃₋₇cycloalkyl, and D is selected from pyridyl and N-oxide of pyridyl.

Other compounds useful in the methods and compositions of the inventionare pharmaceutically-acceptable salts of the compounds in accord withstructural diagram I and tautomers of such a compounds.

Particular compounds of the invention are those wherein E is selectedfrom methyl, phenyl and C₃₋₅cycloalkyl.

More particular compounds of the invention are those according tostructural diagram II:

Still more particular compounds of the invention are those wherein E isselected from methyl, phenyl and C₃₋₅cycloalkyl.

The most particular embodiments of the invention are those exemplarycompounds specifically disclosed herein.

In another aspect the invention provides a method for the treatment ofpain comprising administering a pain-ameliorating effective amount ofany compound according to structural diagram I as heretofore defined.

In particular embodiments the method comprises administeringpain-ameliorating effective amounts of compounds according to structuraldiagram I wherein: E is selected from methyl, phenyl and C₃₋₅cycloalkyl.

In further particular embodiments the method comprises administeringpain-ameliorating effective amounts of compounds according to structuraldiagram II wherein: E is selected from methyl, phenyl andC₃₋₅cycloalkyl.

Yet another aspect of the invention is a pharmaceutical compositioncomprising a pain-ameliorating effective amount of a compound accordingto structural diagram I:

wherein: R¹ is halo; A is CH; E is selected from C₁₋₄alkyl, phenyl andC₃₋₇cycloalkyl; D is selected from pyridyl and N-oxide of pyridyl, ortautomers or pharmaceutically-acceptable salts thereof together with apharmaceutically-acceptable excipient or diluent.

Still more particular embodiments of the invention are those where themethod comprises treatment with an exemplary compound specificallydisclosed herein.

Yet other aspects of the invention are pharmaceutical compositions whichcontain a compound in accord with structural diagram I; the use ofcompounds in accord with structural diagram I for the preparation ofmedicaments and pharmaceutical compositions, and a method comprisingbinding a compound of the invention to the NMDA receptor glycine site ofa warm-blooded animal, such as a human being, so as to beneficiallyinhibit the activity of the NMDA receptor.

DETAILED DESCRIPTION OF THE INVENTION

Compounds of the invention are those within the scope of the genericdescription and particularly those compounds exemplified hereafter.

Suitable pharmaceutically-acceptable salts of compounds of the inventioninclude acid addition salts such as methanesulphonate, fumarate,hydrochloride, hydrobromide, citrate, tris(hydroxymethyl)aminomethane,maleate and salts formed with phosphoric and sulphuric acid. In otherembodiments, suitable salts are base salts such as an alkali metal saltsfor example sodium, alkaline earth metal salts for example calcium ormagnesium, organic amine salts for example triethylamine, morpholine,N-methylpiperidine, N-ethylpiperidine, procaine, dibenzylamine, choline,N,N-dibenzylethylamine or amino acids such as lysine.

Another aspect of the invention is a process for making compounds of theinvention, which process comprises the following steps:

Another aspect of the invention is a process for making compounds of theinvention, which process comprises the following steps:

-   a) Preparing a ketone by reacting a nitrile in the presence of a    Grignard reagent, followed by acidic work-up according to the    following scheme to form said ketone:    where R is an alkyl group;-   b) preparing a Boc-protected hydrazine by reacting a ketone as    prepared in step a), or an aldehyde, according to one of the    procedures shown in the following scheme:-   c) coupling said Boc-protected hydrazine and cyclizing the product    according to the process of the following scheme to form a compound    according to structural diagram I:    wherein:    -   CMC is 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide        metho-p-toluenesulfonate;    -   the “R/H/D” group is the “A-D-E” moiety of structural diagram I,        and    -   throughout the foregoing process R¹ is as defined for structural        diagram I.

To use a compound of the invention or a pharmaceutically-acceptable saltthereof for the therapeutic treatment, which may include prophylactictreatment, of pain in mammals, which may be humans, the compound can beformulated in accordance with standard pharmaceutical practice as apharmaceutical composition.

Suitable pharmaceutical compositions that contain a compound of theinvention may be administered in conventional ways, for example by oral,topical, parenteral, buccal, nasal, vaginal or rectal administration orby inhalation. For these purposes a compound of the invention may beformulated by means known in the art into the form of, for example,tablets, capsules, aqueous or oily solutions, suspensions, emulsions,creams, ointments, gels, nasal sprays, suppositories, finely dividedpowders or aerosols for inhalation, and for parenteral use (includingintravenous, intramuscular or infusion) sterile aqueous or oilysolutions or suspensions or sterile emulsions. A preferred route ofadministration is orally by tablet or capsule.

In addition to a compound of the present invention a pharmaceuticalcomposition of this invention may also contain one or more otherpharmacologically-active agents, or such pharmaceutical composition maybe simultaneously or sequentially co-administered with one or more otherpharmacologically-active agents.

Pharmaceutical compositions of this invention will normally beadministered so that a pain-ameliorating effective daily dose isreceived by the subject. The daily dose may be given in divided doses asnecessary, the precise amount of the compound received and the route ofadministration depending on the weight, age and sex of the patient beingtreated and on the particular disease condition being treated accordingto principles known in the art. A preferred dosage regime is once daily.

A further embodiment of the invention provides a pharmaceuticalcomposition which contains a compound of the structural diagram I asdefined herein or a pharmaceutically-acceptable salt thereof, inassociation with a pharmaceutically-acceptable additive such as anexcipient or carrier.

A yet further embodiment of the invention provide the use of a compoundof the structural diagram I, or a pharmaceutically-acceptable saltthereof, in the manufacture of a medicament useful for binding to theNMDA receptor glycine site in a warm-blooded animal such as a humanbeing.

Still another embodiment of the invention provides a method of binding acompound of the invention to the NMDA receptor-glycine site of awarm-blooded animal, such as a human being, in need of treatment forpain, which method comprises administering to said animal an effectiveamount of a compound of structural diagram I or apharmaceutically-acceptable salt thereof.

Definitions:

When used herein the term “alkyl” includes both straight and branchedchain alkyl groups but references to individual alkyl groups such as“propyl” refer to the straight chain moiety.

When used herein the term “halo” means fluoro, chloro, bromo and iodo.

Generally in the methods, processes and examples described herein:

concentrations were carried out by rotary evaporation in vacuo;

operations were carried out at ambient temperature, that is in the range18-26° C. and under a nitrogen atmosphere;

column chromatography (by the flash procedure) was performed on MerckKieselgel silica (Art. 9385) unless otherwise stated;

yields are given for illustration only and are not necessarily themaximum attainable;

the structure of the end-products of the formula I were generallyconfirmed by NMR and mass spectral techniques, proton magnetic resonancespectra were determined in DMSO-d₆ unless otherwise stated using aVarian Gemini 2000 spectrometer operating at a field strength of 300MHz; chemical shifts are reported in parts per million downfield fromtetramethylsilane as an internal standard (δ scale) and peakmultiplicities are shown thus: s, singlet; bs, broad singlet; d,doublet; AB or dd, doublet of doublets; t, triplet, dt, double oftriplets, m, multiplet; bm, broad multiplet; fast-atom bombardment (FAB)mass spectral data were obtained using a Platform spectrometer (suppliedby Micromass) run in electrospray and, where appropriate, eitherpositive ion data or negative ion data were collected, in thisapplication, (M+H)⁺ is quoted;

intermediates were not generally fully characterized and purity was ingeneral assessed mass spectral (MS) or NMR analysis.

The following abbreviations and definitions when used, have themeanings, as follows:

-   -   CDCl₃ is deuterated chloroform;    -   CMC is 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide        metho-p-toluenesulfonate;    -   DCM is dichloromethane;    -   DCU is dicyclohexyl urea;    -   DHC is 1,3-dicyclohexylcarbodiimide;    -   DMAP is 4-(dimethylamino)pyridine;    -   DMF is N,N-dimethylformamide;    -   DMSO is dimethylsulphoxide;    -   m/s is mass spectroscopy;    -   NMP is N-methylpyrrolidinone;    -   NMR is nuclear magnetic resonance;    -   p.o. is per os;    -   TBF is tetrahydrofuran, and    -   t.i.d. is three times daily.

The examples and tests described herein are intended to illustrate butnot limit the invention.

EXAMPLES Example 1(+/−)-7-Chloro-4-hydroxy-2-(1-cyclopropyl-1-pyrid-2-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione

Cyclopropyl-2-pyridyl ketone.

Dry magnesium powder (0.79 g, 32.6 mmol), which had been previouslycrushed in a mortar with a pestle, was charged into a dry round bottomflask under a nitrogen atmosphere. To this was added diethyl ether (20mL), followed by bromocyclopropane (3.95 g, 2.61 mL, 32.6 mmol). A smallamount of iodine (0.002 g) was added to initiate formation of theGrignard reagent. The reaction was heated to a gentle reflux for a shortperiod of time (˜1 h) until all of the magnesium metal had reacted. Thematerial was allowed to stir at room temperature for 2 hours, and thencooled in an ice bath. To this was added 2-cyanopyridine (2.0 g, 19.2mmol) in diethyl ether (10 mL). The mixture was stirred for 3 hours, andthen carefully quenched with sat. ammonium chloride (3 mL). The reactionwas then acidified with 15% aq. hydrochloric acid (3 mL). The mixturewas stirred for 20 minutes, and then made basic by the addition of 10 Nsodium hydroxide until the pH was approximately 9. To this solution wasadded ethyl acetate (300 mL), and the aqueous layer was extracted withwater (30 mL). The organic layer was washed with brine, dried overNa₂SO₄ and evaporated to dryness giving a yellow oil. This oil waschromatographed (SiO₂, hexanes/ethyl acetate: 75/25) to give the titlecompound as a yellow oil (1.48 g, 52%). ¹H NMR (300 MHz, DMSO-d₆): δ1.10 (m, 4H); 3.46 (m, 1H); 7.70 (m, 1H); 7.96 (t, 1H, J=78. Hz); 8.01(t, 1H, J=7.8 Hz); 8.77 (d, 1H, J=4.8 Hz).

N-1-Aza-2-cyclopropyl-2-(2-pyridyl)vinyl)(tert-butoxy)carboxamide.

To a stirred solution of tert-butylcarbazate (1.32 g, 10.0 mmol) in THF(50 mL) was added cyclopropyl-2-pyridyl ketone (1.48 g, 10.0 mmol),followed by 3 drops of concentrated hydrochloric acid. The reaction wasstirred overnight and the solvent removed in vacuo. The resultant solidwas triturated with hexanes to give the title compound as a white solid(1.90 g, 72%). ¹H NMR (300 MHz, DMSO-d₆): δ 0.86 (m, 4H); 1.45 (s, 9H);2.03 (m, 1H); 7.54 (m, 1H); 8.07 (m, 2H); 8.73 (m, 1H); 13.42 (br s,1H).

(+/−)-(tert-butoxy)-N-[(cyclopropyl-2-pyridylmethyl)amino]carboxamide.

N-1-Aza-2-cyclopropyl-2-(2-pyridyl)vinyl)(tert-butoxy)carboxamide (1.90g, 7.30 mmol) was dissolved in methyl alcohol (90 mL) and placed in aParr shaker bottle. To this was added 10% palladium-on-carbon (300 mg)and the reaction was hydrogenated at 40 psi for 18 h. The catalyst wasfiltered on diatomaceous earth, washed with methyl alcohol (2×300 mL),and the solvents were removed in vacuo to give an oil. This oil waschromatographed (SiO₂, 60/40 hexanes/ethyl acetate and then ethylacetate as eluants) to give the title compound (1.05 g, 54%) as an oil.¹H NMR (300 MHz, DMSO-d₆): δ 0.29 (m, 1H); 0.36 (m, 1H); 0.53 (m, 1H);0.87 (m, 1H); 1.34 (s, 9H); 3.25 (br m, 1H); 4.72 (m, 1H); 7.25 (t, 1H,J=4.8, 8.1 Hz); 7.51 (d, 1H, J=8.1 Hz); 7.75 (t, 1H, J=8.1 Hz); 8.16 (brs, 1H); 8.47 (d, 1H, J=4.8 Hz).

Dimethyl 7-chloro-4-hydroxyquinoline-2,3-dicarboxylate:

A stirred mixture of methyl 2-amino-4-chlorobenzoate (2.50 g, 13.5 mmol)and dimethyl acetylenedicarboxylate (2.05 g, 14.4 mmol) in tert-butanol(22 ml) was refluxed for 7 hours under a nitrogen atmosphere. Afteradding additional dimethyl acetylenedicarboxylate (1.16 g, 8.13 mmol)and refluxing another 2.5 hours, the reaction mixture was allowed tocool to room temperature and potassium tert-butoxide (1.56 g, 13.9 mmol)was added in one portion. A precipitate formed and the resulting mixturewas refluxed for 1.5 hours. The mixture was cooled to room temperatureand filtered to separate the solids, which were washed with tert-butanoland diethyl ether. The solids were dissolved in water and acidified with1 N sulfuric acid to form a precipitate. The resulting mixture wasextracted with DCM and the combined extracts were washed with brine andwater, dried over MgSO₄, filtered and concentrated to give a greensolid. Recrystallization of this material from methanol provided thetitle compound (1.15 g, 47%) as an off-white solid, mp 232-233° C.; MS(Cl):296 (M+H). Analysis for C₁₃H₁₀ClNO₅: Calc'd: C, 52.81; H, 3.41; N,4.74; Found: C, 52.75; H, 3.47; N, 4.69.

3-Carbomethoxy-7-chloro-4-hydroxyquinoline-2-carboxylic acid:

To a stirred suspension of dimethyl7-chloro-4-hydroxyquinoline-2,3-dicarboxylate (1.0 g, 3.38 mmol) inwater (20 mL) was added an aqueous solution of sodium hydroxide (0.27 g,6.75 mmol). Upon addition, the suspension dissolved. The reactionmixture was warmed to 60° C. for 1 hour. After this time the reactionwas cooled to room temperature and acidified with concentratedhydrochloric acid. The product was then extracted into diethyl ether andethyl acetate. The organic extracts were dried over MgSO₄, filtered andconcentrated in vacuo to provide the title compound as a solid (900 mg).This material was purified by recrystallization employing an ethylacetate/hexane co-solvent system to provide the title compound (571 mg,60%) as a white solid mp 296° C. (dec); MS (CI)=238 (M+H). Analysis forC₁₂H₈NO₅Cl.0.45 CH₃CO₂CH₂CH₃.0.10 H₂O: Calc'd: C, 51.30; H, 3.68; N,4.34, Found: C, 51.28; H, 3.62; N, 3.97;¹H NMR 8.22 (d, J=8.7 Hz, 1H),7.92 (d, J=1.8 Hz, 1H), 7.28 (dd, J=8.7, 1.8 Hz, 1H), 3.90 (s, 3H).

3-Carbomethoxy-2-pyrrolidinocarbamide-7-chloro-4-hydroxyquinoline:

To a suspension of3-carbomethoxy-7-chloro-4-hydroxyquinoline-2-carboxylic acid (2.25 g,8.0 mmol) in THF (20 mL) at ambient temperature under a N₂ atmospherewas added DHC (1.65 g, 8.0 mmol) and pyrrolidine (0.596 g, 8.4 mmol).The reaction was stirred room temperature for 15 hours after which timethe by-product urea was removed via filtration. The desired product waspurified via flash column chromatography employing 5% methanol inchloroform to provide the title compound (2.52 g, 94.3%) as a tan solid,mp=215° C.; MS (CI): 335 (M+H). 300 MHz ¹H NMR (DMSO-d₆): δ 8.12 (d,J=8.7 Hz, 1H), 7.60 (d, 1H, J=1.8 Hz), 7.47 (dd, 1H, J=8.8, 2.0 Hz),3.69 (s, 3H), 3.40-3.49 (m, 2H), 3.27-3.33 (m, 2H), 1.80-1.96 (m, 4H).

7-Chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid:

To a suspension of3-carbomethoxy-2-pyrrolidinocarbamide-7-chloro4-hydroxy quinoline (2.52g, 7.5 mmol) in de-ionized water (40 mL) was added dropwise a solution(20 mL) of an aqueous potassium hydroxide (882 mg, 15.75 mmol). Uponcomplete addition, the reaction was warmed to 60° C. After 3 hours, thereaction was filtered to remove a small amount of insoluble material.The filtrate was then acidified to pH=1 which yield a white precipitate.The solid was isolated by vacuum filtration, washed with water, anddried at 30° C. in vacuo for 16 hours. This provided the title compound(1.5 g, 64%) as a white solid, mp=225-8° C.; MS (CI): 321 (M+H). 300 MHz¹H NMR (DMSO-d₆): δ 8.28 (d, J=8.8 Hz, 1H), 7.77 (s, 1H), 7.64 (d, 1H,J=8.7), 3.52-3.57 (m, 2H), 3.17-3.19 (m, 2H), 1.83-1.98 (m, 4H).

(+/−)-N-[(tert-Butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopropyl-2-pyridylmethyl)carboxamide.

To a stirred slurry of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid(1.27 g, 3.99 mmol) in THF (60 mL) was added CMC (1.93 g, 4.56 mmol) andthe reaction was stirred for five minutes. To this mixture was added, bydropwise addition, a solution of(+/−)-(tert-butoxy)-N-[(cyclopropyl-2-pyridylmethyl)amino]carboxamide(1.00 g, 3.80 mmol) and DMAP (0.040 g, 0.32 mmol) in THF (20 mL), andthe mixture was stirred at room temperature for 1 hour. The mixture wasthen refluxed overnight. The solution was filtered and the insolubleswashed with DCM (2×150 mL). The mother liquor was collected andconcentrated to dryness. The resultant solid was subjected tochromatography (SiO₂, 95/5 chloroform/methyl alcohol) to give the titlecompound as a yellow foam (2.08 g, 96%). This material was used as is inthe next reaction.

(+/−)-7-Chloro4-hydroxy-2-(1-cyclopropyl-1-pyrid-2-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione.

To a stirred solution of(+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopropyl-2-pyridylmethyl)carboxamide:(2.08 g, 3.68 mmol) in THF (40 mL) was added methanesulfonic acid (9 mL)and the reaction was stirred overnight. The volatiles were removed invacuo and to the residual oil was added diethyl ether (80 mL). Themixture was stirred for 10 minutes and then allowed to settle into twolayers, an etheral layer and layer of brown oil. The ether was decantedaway and the brown oil was dissolved in water (100 mL). This solutionwas cooled in an ice bath and to this was added sodium chloride (sat.aqueous, 10 mL), followed by solid sodium chloride (5 g). After a shorttime, a precipitate formed. This precipitate was collected by vacuumfiltration and washed with diethyl ether (2×100 mL). The residue wasthen sonicated in 20 mL of 4/1 diethyl etherimethyl alcohol for fifteenminutes. The material was filtered, washed with diethyl ether and driedin vacuo to give the title compound (0.574 g, 32%) as an off-whitepowder (m.p.>280° C.). ¹H NMR (300 MHz, DMSO-d₆): δ 0.53 (m, 1H); 0.70(m, 2H); 0.86 (m, 1H); 1.74 (m, 1H); 5.49 (m, 1H); 7.45 (d, 1H, J=8.7Hz); 7.71 (m, 1H); 7.97 (m, 2H); 8.09 (m, 1H); 8.28 (m, 1H); 8.73 (m,1H); 11.97 (br s, 1H); 12.75 (br s, 1H). Calc'd. forC₂₀H₁₅ClN₄O₃.1.0NaCl.1.0H₂O: C, 50.58; H, 3.69; N, 11.79; Found: C,50.67; H, 3.71; N, 11.84.

Example 2(+/−)-7-Chloro-4-hydroxy-2-(1-cyclopropyl-1-pyrid-4-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionehydrochloride

Cyclopropyl-4-pyridyl ketone.

Dry magnesium powder (0.760 g, 31.2 mmol), which had been previouslycrushed in a mortar with a pestle, was charged into a dry round bottomflask under a nitrogen atmosphere. To this was added diethyl ether (20mL), followed by bromocyclopropane (3.77 g, 2.49 mL, 31.2 mmol). A smallamount of iodine (0.002 g) was added to initiate formation of theGrignard reagent. The reaction was heated to a gentle reflux for a shortperiod of time (˜1 hour) until all of the magnesium metal had reacted.The flask was then cooled in an ice bath and to this was added4-cyanopyridine (1.62 g, 15.6 mmol). The mixture was stirred for 3hours, and then carefully quenched with sat. ammonium chloride (3 mL).The reaction was then acidified with 15% aq. hydrochloric acid (3 ML).The mixture was stirred for 20 minutes, and then made basic by theaddition of 10 N sodium hydroxide until the pH was approximately 9. Tothis solution was added ethyl acetate (300 mL), and the aqueous layerwas extracted with water (30 mL). The organic layer was washed withbrine, dried over Na₂SO₄ and evaporated to dryness giving the titlecompound as a yellow oil (1.0 g, 43%). ¹H NMR (300 MHz, DMSO-d₆): δ 1.15(m, 4H); 2.93 (s, 1H); 7.91 (d, 1H; J=6.0 Hz); 8.83 (d, 1H, J=6.0 Hz).

N-1-Aza-2-cyclopropyl-2-(4-pyridyl)vinyl)(tert-butoxy)carboxamide.

To a stirred solution of tert-butylcarbazate (0.95 g, 7.21 mmol) in THF(50 mL) was added cyclopropyl-4-pyridyl ketone (1.06 g, 7.21 mmol),followed by 3 drops of concentrated hydrochloric acid. The reaction wasstirred overnight and the solvent removed in vacuo. The resultant solidwas chromatographed (SiO₂, ethyl acetate as eluant, R_(f)=0.25) to givethe title compound as a white solid (0.98 g, 52%). ¹H NMR (300 MHz,DMSO-d₆): δ 0.40 (m, 2H); 1.07 (m, 2H); 1.49 (s, 9H); 1.76 (m, 1H); 7.60(d, 2H, J=4.8 Hz); 8.56 (d, 1H, J=4.8Hz).

(+/−)-(tert-Butoxy)-N-[(cyclopropyl-4-pyridylmethyl)amino]carboxamide.

N-1-Aza-2-cyclopropyl-2-(4-pyridyl)vinyl)(tert-butoxy)carboxamide (0.98g, 3.75 mmol) was dissolved in methyl alcohol (90 mL) and placed in aParr shaker bottle. To this was added 10% palladium-on-carbon (500 mg)and the reaction was hydrogenated at 40 psi for 18 hours. The catalystwas filtered on diatomaceous earth, washed with methyl alcohol (2×300mL), and the solvents were removed in vacuo to give an oil. The materialwas used as is with no further purification (0.77 g, 78%). ¹H NMR (300MHz, DMSO-d₆): δ 0.34 (m, 2H); 0.58 (m, 1H); 0.80 (m, 1H); 1.33 (s, 9H);1.75 (m, 1H); 3.33 (br s, 1H); 4.75 (br s, 1H); 7.38 (d, 2H, J=6.0 Hz);8.20 (br s, 1H); 8.45 (d, 2H, J=6.0 Hz).

(+/−)-N-[(tert-Butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopropyl-4-pyridylmethyl)carboxamide.

To a stirred slurry of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid,Example 1, (0.99 g, 3.10 mmol) in THF (50 mL) was added CMC (1.48 g,3.54 mmol) and the reaction was stirred for five minutes. To thismixture was added, via dropwise addition, a solution of(+/−)-(tert-butoxy)-N-[(cyclopropyl-4-pyridylmethyl)amino]carboxamide(0.77 g, 2.95 mmol) and DMAP (0.05 g, 0.40 mmol) in THF (10 mL), and themixture was stirred at room temperature for 1 hour. The mixture was thenrefluxed overnight. The solution was filtered hot and the insolubleswashed with DCM (2×150 mL). The mother liquor was collected andconcentrated to dryness. The resultant solid was subjected tochromatography (SiO₂, 95/5 chloroform/methyl alcohol) to give the titlecompound as a yellow foam. This foam was then triturated with diethylether to give the title compound (1.2 g, 68%) as a yellow solid.

(+/−)-7-Chloro-4-hydroxy-2-(1-cyclopropyl-1-pyrid-4-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionehydrochloride.

To a stirred solution of(+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopropyl-4-pyridylmethyl)carboxamide(1.2 g 2.12 mmol) in THF (40 mL) was added methanesulfonic acid (5 mL)and the reaction was stirred overnight. The volatiles were removed invacuo and to the residual oil was added diethyl ether (80 mL). Themixture was stirred for 10 minutes and then allowed to settle into twolayers, an etheral layer and layer of brown oil. The ether was decantedaway and to the brown oil was added water (18 mL). After a short time, aprecipitate formed and was discarded. To the remaining filtrate wasadded solid sodium chloride until the liquid was saturated. Thismaterial was left to stand until a precipitate formed. This precipitatewas collected by vacuum filtration and washed with diethyl ether (2×100mL). The solids were then sonicated in water (8 mL), filtered andsonicated in 10 mL of 4/1 diethyl ether/methyl alcohol for fifteenminutes. The material was filtered, washed with diethyl ether and driedin vacuo to give the title compound (0.182 g, 19%) as an off-whitepowder (m.p>280° C.). ¹H NMR (300 MHz, DMSO-d₆): δ 0.46 (m, 1H); 0.65(m, 1H); 0.85 (m, 2H); 1.67 (m, 1H); 5.37 (d, 1H, J=10.2 Hz); 7.45 (dd,1H, J=2.1, 8.7 Hz); 7.93 (d, 2H, J=5.4 Hz); 8.05 (d, 1H, J=2.1 Hz); 8.14(d, 2H, J=5.4 Hz); 8.81 (d, 2H, J=5.4 Hz); 12.04 (br s, 1H); 12.75 (brs, 1H). Calc'd. for C₂₀H₁₅ClN₄O₃.1.0HCl.1.2H₂O: C, 53.04; H, 4.09; N,12.37; Found: C, 53.22; H, 4.01; N, 11.93.

Example 3(+/−)-7-Chloro-4-hydroxy-2-(1-cyclopentyl-1-pyrid-2-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionehydrochloride

Cyclopentyl-2-pyridyl ketone.

Prepared in an analogous fashion to cyclopropyl-2-pyridyl ketone,Example 1, (45% as an oil). ¹H NMR (300 MHz, DMSO-d₆): δ 1.66 (m, 6H);1.84 (m, 2H); 4.17 (m, 1H); 7.66 (m, 1H); 7.98 (m, 2H); 8.74 (d, 1H,J=4.8 Hz).

N-1-Aza-2-cyclopentyl-2-(2-pyridyl)vinyl)(tert-butoxy)carboxamide.

To a stirred solution of tert-butylcarbazate (2.43 g, 13.8 mmol) in THF(50 mL) was added cyclopentyl-2-pyridyl ketone (1.83 g, 13.8 mmol),followed by 3 drops of concentrated hydrochloric acid. The reaction wasstirred overnight and the solvent removed ini vacuo. The resultant solidwas triturated with hexanes to give the title compound as a white solid(3.35 g, 84%). ¹H NMR (300 MHz, DMSO-d₆): δ 1.45 (s, 9H); 1.66 (m, 6H);1.84 (m, 2H); 3.37 (m, 1H); 7.51 (t, 1H, J=4.8, 7.8 Hz); 7.80 (d, 1H,J=8.1 Hz); 8.02 (dd, 1H, J=1.5, 7.8 Hz); 8.74 (d, 1H, J=4.8 Hz).

(+/−)-(tert-Butoxy)-N-[(cyclopentyl-2-2pyridylmethyl)amino]carboxamide.

N-1-Aza-2-cyclopentyl-2-(2-pyridyl)vinyl)(tert-butoxy)carboxamide (2.00g, 6.9 mmol) was dissolved in methyl alcohol (90 mL) and placed in aParr shaker bottle. To this was added 10% palladium-on-carbon (800 mg)and the reaction was hydrogenated at 40 psi for 18 hours. The catalystwas filtered on diatomaceous earth, washed with methyl alcohol (2×300mL), and the solvents were removed in vacuo to give the title compound(1.92 g, 96%) as an oil. ¹H NMR (300 MHz, DMSO-d₆): δ 1.17 (m, 2H); 1.33(s, 9H); 1.52 (m, 6H); 1.81 (m, 1H); 2.04 (m, 1H); 4.10 (d, 1H, J=4.5Hz); 4.61 (m, 1H); 7.21 (d, 1H, J=4.8, 7.8 Hz); 7.43 (d, 1H, J=7.8 Hz);7.73 (dd, 1H, J=1.5, 7.8 Hz); 8.068 (br s, 1H); 8.44 (d, 1H, J=4.8 Hz).

(+/−)-N-[(tert-Butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopentyl-2-pyridylmethyl)carboxamide.

To a stirred slurry of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid,Example 1, (2.21 g, 6.89 mmol) in THF (80 mL) was added CMC (3.34 g,7.90 mmol) and the reaction was stirred for five minutes. To thismixture was added, by dropwise addition, a solution of(+/−)-(tert-butoxy)-N-[(cyclopentyl-2-pyridylmethyl)amino]carboxamide(1.92 g, 6.59 mmol) and DMAP (0.15 g, 1.22 mmol) in THF (10 mL), and themixture was stirred at room temperature for 1 hours. The mixture wasthen refluxed overnight. The solution was filtered hot and theinsolubles washed with DCM (2×150 mL). The mother liquor was collectedand concentrated to dryness. The resultant solid was subjected tochromatography (SiO₂, 95/5 chloroform/methyl alcohol) to give the titlecompound as a yellow foam. This foam was then triturated with diethylether to give the title compound (3.14 g, 80%) as a yellow solid.

(+/−)-7-Chloro-4-hydroxy-2-(1-cyclopentyl-1-pyrid-2-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionehydrochloride.

To a stirred solution of (+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(cyclopentyl-2-pyridylmethyl)carboxamide(3.14 g, 5.29 mmol) in THF (100 mL) was added methanesulfonic acid (13mL) and the reaction was stirred overnight. The volatiles were removedin vacuo and to the residual oil was added diethyl ether (200 mL). Themixture was stirred for 10 minutes and then allowed to settle into twolayers, an etheral layer and layer of brown oil. The ether was decantedaway and to the brown oil was added water (20 mL), followed by sodiumchloride (sat. aqueous, 25 mL). After a short time, a precipitate formedand was collected by vacuum filtration. The precipitate was sonicated inwater (10 mL), filtered and sonicated in 20 mL of 4/1 diethylether/methyl alcohol for fifteen minutes. The material was filtered,washed with diethyl ether and dried in vacuo to give the title compound(0.632 g, 24%) as an off-white powder (m.p>220-225° C.). ¹H NMR (300MHz, DMSO-d₆): δ 1.27 (m, 2H); 1.55 (m, 3H); 1.70 (m, 3H); 3.02 (m, 1H);5.95 (d, 1H, J=11.1 Hz); 7.45 (dd, 1H, J=2.1, 8.7 Hz); 7.67 (t, 1H,J=6.0 Hz); 7.84 (d, 1H, J=8.1 Hz); 8.03 (d, 1H, J=2.1 Hz); 8.14 (d, 2H,J=5.4 Hz); 8.21 (t, 1H, J=7.8 Hz); 8.72 (d, 1H, J=4.8 Hz); 12.04 (br s,1H); 12.70 (br s, 1H). Calc'd. for C₂₂H₁₉ClN₄O₃.1.0 HCl.1.5H₂O: C,54.33; H, 4.76; N, 11.51; Found: C, 54.03; H, 4.40; N, 11.39.

Example 4(+/−)-7-Chloro-4-hydroxy-2-(1-phenyl-1-pyrid-4-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione

N-1-Aza-2-Phenyl-2-(4-pyridyl)vinyl)(tert-butoxy)carboxamide.

A solution of 4-benzoylpyridine (5.0 g, 27.3 mmoles),tert-butylcarbazate (3.61 g, 27.3 mmoles), and 3 drops of concentratedhydrochloric acid in THF was stirred at room temperature for 18 hours.The reaction was concentrated under reduced pressure to give a yellowoil. The oil was purified by trituration with ether/hexane (1/1) toobtain the title compound as a white solid (5.43 g, 67% yield). ¹H NMR(300 MHz, DMSO-d₆): δ 1.44 (s, 9H), 7.32 (m, 3H); 7.38 (m, 3H, ); 7.58(m, 1H;8.57 (d, 1H, J=1.5 Hz ); 8.74 (d, 1H, J=1.5 Hz); 9.15 (s, 1H);9.48 (s, 1H).

(+/−)-(tert-Butoxy)-N-[(phenyl-4-pyridylmethyl)amino]carboxamide.

A mixture of 10% palladium on carbon (0.55 g) andN-1-aza-2-phenyl-2-(4-pyridyl)vinyl)(tert-butoxy)carboxamide (2.06 g,6.93 mmol) in methanol (200 mL). was hydrogenated (50 psi) at roomtemperature for 18 hour. The reaction was filtered through diatomaceousearth and the filtrate evaporated under reduced pressure to give thetitle compound as a gold oil (2.08 g, quantitative recovery). ¹H NMR(300 MHz, DMSO-d₆): δ 1.36 (s, 9H); 5.10 (s, 1H); 5.27 (s, 1H, ); 7.31(m, 3H); 7.42 (m, 4H); 8.36 (s, 1H); 8.75 (d, 2H, J=4 Hz).

(+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl)]-N-(phenyl-4-pyridylmethyl)carboxamide.

A mixture of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid,Example 1, (3.60 g, 11.2 mmol),(+/−)-(tert-butoxy)-N-[(phenyl-4-pyridylmethyl)amino]carboxamide (3.37g, 11.2 mmol), and cyclohexyl-3-(2-morpholinoethyl)carbodiimidemetho-p-toluenesulfonate (7.20 g, 17.0 mmol) in THF (350 mL, dry) wasrefluxed for 18 hours. The reaction was filtered and the filtrateconcentrated under reduced pressure. The residue was taken up in DCM(300 mL), washed with water (2×200 mL), and dried over MgSO₄.Concentration of the organic layer in vacuo provided a gold oil. Thetitle compound was purified by silica gel column chromatography usingmethanol/chloroform (5/95) as eluant. The title compound was obtain asan tan solid (3.97 g, 59%). This material was used in the followingreaction. MS (+CI) m/z 600/602.

(+/−)-7-Chloro-4-hydroxy-2-(1-phenyl-1-pyrid-4-yl)methyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione.

A solution ofN-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl)]-N-(phenyl-4-pyridylmethyl)carboxamide(3.94 g, 6.54 mmol), methanesulfonic acid (5 mL, 7.41 g, 77 mmol), andTHF (100 mL) were stirred at room temperature for 48 hours. The reactionwas concentrated in vacuo to a gold oil. The oil was taken up in waterand filtered to remove insolubles. The filtrate was neutralized withsodium hydroxide (5 N aqueous, pH=7), and the resulting solid filteredand sonicated in methyl alcohol/diethyl ether (1/1, 20 ml). The mixturewas filtered and the collected solids dried to give the title compound(0.30 g, 6.0% yield). ¹H NMR (300 MHz, DMSO-d₆): δ 2.32 (s, 3H,methanesulfonic acid), 7.17 (d, 2H, J=6 Hz); 7.40 (m, 8H); 8.07 (d, 1H,J=4.2 Hz); 8.17 (d, 1H, J=8.7 Hz); 8.48 (d, 2H, J=6 Hz); 12.73(br s,1H). Calc'd. for C₂₃H₁₅ClN₄O₃.3.0CH₃SO₃Na.0.25H₂O: C, 39.13; H, 3.16; N,6.96. Found: C, 39.29; H, 3.18; N, 7.00.

Example 5(+/−)-7-Chloro-4-hydroxy-2-(1-(3-methylpyrid-4-yl))ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione

4-Cyano-3-methylpyridine.

To a cooled (0° C.) round bottom flask charged with 3-picoline-N-oxide(4.53 g, 42 mmol) was added dimethylsulfate (5.8 g, 47 mmol). Themixture was stirred for 18 hours and allowed to warm to roomtemperature. To this mixture was added potassium cyanide (3.25 g, 50mmol) dissolved in ethyl alcohol/water (75 mL, 1:1). The reaction wasstirred at room temperature for 18 hours, concentrated to half thevolume and diluted with chloroform (100 mL). The organic layer wasseparated and washed with sodium chloride (sat. 25 mL) and dried overNa₂SO₄. The solvent was removed and the residual material waschromatographed (SiO₂, hexanes/ethyl acetate) to give the title compoundas off-white crystals (2.01 g, 41%). ¹H NMR (300 MHz, DMSO-d₆): δ 2.55(s, 3H); 7.47 (d, 1H); J=5.1 Hz); 8.60 (d, 1H, J=5.1 Hz); 8.68 (s, 1H).

The intermediate pyridylketone was made using commercially availablemethylmagnesium bromide (1.4 M in toluene/THF) and the nitrile describedabove using the method previously described in Example 1. Conversion to(+/−)-7-chloro-4-hydroxy-2-(1-methylpyridy-4-yl)methyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionewas likewise accomplished in a manner analogous to Example 1 to give anoff-white powder. ¹H NMR (300 MHz, DMSO): δ 1.65 (d, 3H, J=6.9 Hz); 2.34(s, 3H, CH₃SO₃H); 2.53 (s, 3H); 6.14 (q, 1H, J=6.9 Hz); 7.42 (dd, 1H,J=1.8, 8.7 Hz); 7.87 (d, 1H, J=6.3 Hz); 8.04 (d, 1H, J=1.8 Hz); 8.13 (d,1H, J=8.7 Hz); 8.73 (d, 1H, J=6.3 Hz); 8.80 (s, 1H); 12.02 (br s, 1H);12.73 (br s, 1H).

Example 6(+/−)-7-Chloro-4-hydroxy-2-(1-methyl(6-methylpyrid-2-yl)methyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate

2-Cyano-6-methylpyridine.

2-Cyano-6-methyypyridine was made using 2-picoline-N-oxide as thestarting material in a manner analogous to the procedure described inExample 5: ¹H NMR (300 MHz, DMSO-d₆): δ 2.61 (s, 3H); 7.38 (d, 1H, J=7.8Hz): 7.49 (d, 1H, J=7.8 Hz); 7.72 (t, 1H, J=7.8 Hz).

The title compound, an off-white powder, was prepared in a manneranalogous to Example 5. ¹H NMR (300 MHz, DMSO-d₆-TFA shake): δ 1.78 (d,3H, J=6.9 Hz); 2.79 (s, 3H); 6.44 (q, 1H, J=6.9 Hz); 7.42 (dd, 1H,J=2.1, 8.7 Hz); 7.78 (s, 1H); 7.96 (m, 3H); 8.56 (t, 1H, J=8.1 Hz).Calc'd. for C₁₉H₁₅ClN₄O₃.1.1 CH₃SO₃H.0.45 H₂O: C, 48.61; H, 4.12; N,11.28; Found: C, 48.72; H, 4.18; N, 11.19.

In the conversion of 2-picoline-N-oxide to 2-cyano-6-methylpyridine, thereaction gave rise to a side-product, 4-cyano-2-methylpyridine, whichwas separated from the title compound by column chromatography (SiO₂,hexane/ethyl acetate 90:10 to 80:20 gradient). 4-cyano-2-methylpyridine:¹H NMR (300 MHz, DMSO-d₆): δ 2.64 (s, 3H); 7.34 (d, 1H, J=5.1 Hz); 7.40(s, 1H); 8.68 (d, 1H, J=5.1 Hz). This material was used in Example 8,below.

Example 7(+/−)-7-Chloro-4-hydroxy-2-(1-methyl(3-methylpyrid-2-yl)methyl)-1,2,5,10-tetrahydropydridazino[4,5-b]quinoline-1,10-dionemethanesulfonate

The title compound was made in a manner analogous to the proceduredescribed in Example 5, using 2-cyano-3-methylpyridine andmethylmagnesium bromide (1.4 M in toluene/THF) as starting materials togive the title compound as an off-white powder. ¹H NMR (300 MHz,DMSO-d₆-tfa shake): δ 1.82 (d, 3H, J=6.9 Hz); 2.48 (s, 3H); 6.21 (q, 1H,J=6.9 Hz); 7.45 (dd, 1H, J=1.8, 8.7 Hz); 7.96 (dd, 1H, J=5.7, 7.5 Hz);8.06 (d, 1H, J=1.8 Hz); 8.16 (d, 1H, J=8.7 Hz); 8.48 (d, 1H, J=7.5 Hz);8.73 (d, 1H, J=4.8 Hz). Calc'd. for C₁₉H₁₅ClN₄O₃.4 CH₃SO₃H.0.2H₂O: C,35.84; H, 4.11; N, 7.27; Found: C, 35.47; H, 3.73; N, 7.36.

Example 8(+/−)-7-Chloro-4-hydroxy-2-(1-methyl(2-methylpyrid-4-yl)methyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate

The title compound was made by analogy to the procedure described inExample 1, using 4-cyano-2-methylpyridine (described in Example 6) andmethylmagnesium bromide (1.4 M in toluene/THF) as starting materials togive the title compound as an off-white powder. ¹H NMR (300 MHz,DMSO-d₆): δ 1.72 (d, 3H, J=6.9 Hz); 2.32 (s, 3H, CH ³ SO₃H); 2.67 (s,3H); 6.21 (q, 1H, J=6.9 Hz); 7.45 (dd, 1H, J=1.8, 8.7 Hz); 7.76 (d, 1H,J=6.0 Hz); 7.79 (s, 1H); 8.06 (d, 1H, J=1.8 Hz); 8.15 (d, 1H, J=8.7 Hz);8.66 (d, 1H, J=6.0 Hz); 12.04 (br s, 1H); 12.70 (br s, 1H). Calc'd. forC₁₉H₁₅ClN₄ _(O) ₃.1.35CH₃SO₃H.1.05H₂O: C, 45.99; H, 4.27; N, 10.54;Found: C, 46.00; H. 4.21; N, 10.60.

Example 9(+/−)-7-Chloro-4-hydroxy-2-(1-methyl(3-trifluoromethylpyrid-2-yl)methyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione

2-Cyano-3-trifluromethylpyridine

To a stirred solution of 2-chloro-3-trifluoromethylpyridine (5.00 g,27.5 mmol) in DMSO (20 mL) was added potassium cyanide (2.15 g, 33.0mmol) and the mixture heated to 100° C. for 18 hours. The mixture wasdiluted with ethyl acetate and extracted with water (80 mL). The organiclayer was separated and evaporated to give a brown residue. The materialwas chromatographed (SiO₂, hexanes/ethyl acetate gradient 85/15 to80/20) to give the intermediate nitrile as an oil (0.8 g, 17%). ¹H NMR(300 MHz, DMSO-d₆): δ 7.73 (dd, 1H, J=8.1, 4.5 Hz); 8.15 (d, 1H, J=8.1Hz); 8.92 (d, 1H, J=4.5 Hz).

The intermediate pyridylketone was made using commercially availablemethyl magnesium bromide (1.4 M in toluene/THF) and the nitriledescribed above using the method previously described in Example 5.Conversion to(+/−)-7-chloro-4-hydroxy-2-(1-methyl(3-trifluromethylpyridy-2-yl)methyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionewas likewise accomplished by a process analogous to that in Example 5 togive an off white powder. ¹H NMR (300 MHz, DMSO-d₆): δ 1.67 (d, 3H,J=6.6 Hz); 6.181 (q, 1H, J=6.6 Hz); 7.42 (d, 1H, J=8.7 Hz); 7.53 (dd,1H, J=5.1, 7.8 Hz); 8.03 (d, 1H, J=2.1 Hz); 8.14 (d, 1H, J=8.7 Hz); 8.16(d, 1H, J=7.8 Hz); 8.76 (d, 1H, J=4.5 Hz); 11.92 (br s, 1H); 12.40 (brs, 1H).

Example 10(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-2-yl-N-oxide)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione

N-(1-Aza-2-(2-pyridyl)prop)-1-enyl)(tert-butoxy)carboxarride.

To a stirred solution of tert-butylcarbazate (2.18 g, 16.5 mmol) in THF(40 mL) was added 2-acetylpyridine (2.00 g, 16.5 mmol), followed by 3drops of concentrated hydrochloric acid. After 1 h, the reaction turnedcloudy, and the solvent was removed in vacuo. The resultant solid wastriturated with hexanes and filtered to give the title compound as awhite solid (3.12 g, 80%). ¹H NMR (300 MHz, DMSO-d₆): δ 1.49 (s, 9H);7.38 (dd, 1H, J=4.8, 6.7 Hz); 7.94 (m, 1H); 7.99 (d, 1H, J=7.5 Hz); 8.58(d, 1H, J=4.2 Hz); 10.04 (s, 1H).

(+/−)-(tert-Butoxy)-N-[(2-pyridylethyl)amino]carboxamide.

N-(1-Aza-2-(2-pyridyl)prop-1-enyl)(tert-butoxy)carboxamide (2.0 g, 8.5mmol) was dissolved in methyl alcohol (80 mL) and placed in a Parrshaker bottle. To this was added 10% palladium-on-carbon (850 mg) andthe reaction was hydrogenated at 40 psi for 24 h. The mixture wasfiltered through diatomaceous earth, which was washed with methylalcohol (3×100 mL). The combined filtrate and washes were concentratedin vacuo. The resultant oil (ca. 1.8 g) was used in the followingreaction without further purification. ¹H NMR (300 MHz, DMSO-d₆): δ 1.19(d, 3H, J=6.6 Hz); 1.33 (s, 9H); 4.11 (m, 1H); 4.79 (m, 1H); 7.22 (m,1H); 7.49 (d, 1H, J=7.8 Hz); 8.22 (m, 1H, J=1.5, 2.4 Hz); 8.49 (d, 1H,J=4.2 Hz).

(+/−)-N-[(tert-Butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(2-pyridylethyl)carboxamide.

To a stirred slurry of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid,Example 1, (2.43 g, 7.57 mmol) in THF (100 mL) was added CMC (3.69 g,8.72 mmol) and the reaction was stirred for five minutes. To thismixture was added dropwise a solution of(+/−)-(tert-butoxy)-N-[(2-pyridylethyl)amino]carboxamide (1.8 g, 7.59mmol) and DMAP (0.160 g, 1.30 mmol) in THF (20 mL). The mixture wasstirred at room temperature for 45 minutes and then refluxed overnight.The cooled solution was filtered and the collected insolubles washedwith DCM (2×150 mL). The combined filtrate and washes were concentratedin vacuo to dryness. The resultant yellow foam was subjected tochromatography (silica gel, 95/5 chloroform/methyl alcohol) to give thetitle compound as a yellow foam (3.3 g, 81%).

(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-2-yl)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate.

To a stirred solution of(+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(2-pyridylethyl)carboxamide(3.0 g, 5.56 mmol) in THF (100 mL) was added methanesulfonic acid (15mL) and the reaction was stirred overnight. The volatiles were removedin vacuo and the resultant oil was poured on to crushed ice. A fineprecipitate formed which was filtered to give an orange solid. Thismaterial was washed with diethyl ether, and then sonicated in 20 mL of1/1 diethyl ether/methyl alcohol for fifteen minutes and filtered. Thecollected solids were sonicated again in 85 mL of the same solventsystem for an additional fifteen minutes. The insoluble materials werecollected, washed with the same solvent system and dried at 55 ° C. for12 h to give the title compound (1.29 g, 48%) as an off-white powder(m.p.>290 ° C). ¹H NMR (300 MHz, DMSO-d₆): δ 1.77 (d, 3H, J=6.9 Hz);2.34 (s, 3H, CH₃SO₃H); 6.38 (q, 1H, J=6.9 Hz); 7.44 (dd, 1H, J=1.5, 8.7Hz); 7.82 (m, 2H); 7.90 (d, 1H, J=8.1 Hz); 8.04 (d, 1H, J=8.7 Hz); 8.38(app t, 1H, J=7.5 Hz); 8.82 (d, 1H, J=5.1 Hz); 11.98 (s, 1H); 12.80 (s,1H). Calc'd. for C₁₈H₁₃ClN₄O₃.CH₃SO₃H.H₂O: C, 47.25; H, 3.96; N, 11.60.Found: C, 47.26; H, 3.67; N, 11.50.

(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-2-yl-N-oxide)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione.

To a stirred suspension of(+/−)-7-chloro-4-hydroxy-2-(1-pyrid-2-yl)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate (0.420 g, 0.86 mmol) in methyl alcohol (1.0 mL) wasadded choline hydroxide (0.49 mL, 45% in MeOH, 1.73 mmol) and thesolution stirred until all had dissolved. To this was added3-chloroperoxybenzoic acid (0.316 g, 57-86% purity, 1.04 mmol -1.57mmol) and the solution stirred 24 hours. An additional portion of3-chloroperoxybenzoic acid (0.08 g, 57-86% purity, 0.2 mmol-0.3 mmol)was added and the reaction stirred 2 days. The solid was then filtered,washed with methyl alcohol. The material was triturated in diethylether, filtered and dried in vacuo (200 mtorr, 60° C., 2 hours) to givethe title compound (0.157 g, 47%) as a yellow solid. ¹H NMR (300 MHz,DMSO-d₆): δ 1.65 (d, 3H, J=6.9 Hz) 6.35 (d, 1H, J=6.9 Hz); 7.32 (m, 3H);7.42 (d, 1H, J=8.7 Hz); 8.03 (s, 1H); 8.14 (d, 1H, J=8.7 Hz); 8.24 (d,1H, J=5.7 Hz); 11.91 (br s, 1H); 12.50 (br s, 1H). MS (+CI) m/z 385/387.

Example 11(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-4-yl-N-oxide)ethyl-1,2,5,10-tetrahydrpyridazino[4,5-b]quinoline-1,10-dione

N-(1-aza-2-(4-pyridyl)prop-1-enyl)(tert-butoxy)carboxamide.

To a stirred solution of tert-butylcarbazate (2.18 g, 16.5 mmol) in THF(40 mL) was added 4-acetylpyridine (2.00 g, 16.5 mmol), followed by 3drops of concentrated hydrochloric acid. After 1 h, the reaction turnedcloudy, and the solvent was removed in vacuo. The resultant solid wastriturated with hexanes and filtered to give the title compound as awhite solid (3.88 g, 100%). ¹H NMR (300 MHz, DMSO-d₆): δ 1.47 (s, 9H);7.55 (dd, 2H, J=1.5, 4.5 Hz); 7.97 (s, 1H); 7.99 (d, 2H, J=4.5 Hz);11.22 (s, 1H).

(+/−)-(tert-Butoxy)-N-[(4-pyridylethyl)amino]carboxamide.

N-(1-Aza-2-(4-pyridyl)prop-1-enyl)(tert-butoxy)carboxamide (2.0 g, 8.51mmol) was dissolved in ethyl alcohol (90 mL) and placed in a Parr shakerbottle. To this was added 10% palladium-on-carbon (500 mg) and thereaction was hydrogenated at 40 psi for 24 h. The mixture was filteredthrough diatomaceous earth, which was washed with methyl alcohol (3×100mL). The combined filtrate and washes were concentrated in vacuo. Theresultant material was triturated with 90/10 hexanes/DCM to give thetitle compound as a white solid (1.34 g, 66%). ¹H NMR (300 MHz,DMSO-d₆): δ 1.19 (d, 3H, J=6.6 Hz); 1.34 (s, 9H); 4.09 (m, 1H); 4.86 (m,1H); 7.35 (d, 2H, J=5.7 Hz); 8.21 (s, 1H); 8.47 (d, 2H, J=5.7 Hz).

(+/−)-N-[(tert-Butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(4-pyridylethyl)carboxamide.

To a stirred slurry of7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)hydroquinoline-3-carboxylic acid,Example 1, (1.66 g, 5.17 mmol) in THF (40 mL) was added CMC (2.82 g,6.72 mmol) and the reaction was stirred for five minutes. To thismixture was added dropwise a solution of(+/−)-(tert-butoxy)-N-[(4-pyridylethyl)amino]carboxamide (1.3 g, 5.69mmol) and DMAP (0.080 g, 0.65 mmol) in THF (20 mL). After stirring thereaction mixture at room temperature for 2 hours, an additional portionof CMC (0.500 g) was added and the mixture was refluxed overnight. Thesolution was cooled to 50° C., another portion of diimide added (0.500g) and the mixture was refluxed for 3 h. The cooled reaction mixture wasfiltered and the collected insolubles washed with DCM (2×150 mL). Thecombined filtrate and washes were concentrated in vacuo to dryness. Theresultant yellow foam was subjected to chromatography (silica gel, 92/8chloroform/methyl alcohol) to give the title compound as a yellow foam(2.04 g, 73%).

(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-4-yl)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate.

To a stirred solution of (+/−)-N-[(tert-butoxy)carbonylamino][7-chloro-4-oxo-2-(pyrrolidinylcarbonyl)(3-hydroquinolyl]-N-(4-pyridylethyl)carboxamide(2.04 g, 3.8 mmol) in THF (100 mL) was added methanesulfonic acid (13.5mL) and the reaction was stirred overnight. The volatiles were removedin vacuo and the resultant oil was poured on to crushed ice. A fineprecipitate formed which was filtered to give an orange solid. Thismaterial was washed with diethyl ether, and then sonicated in 40 mL of1/1 diethyl ether/methyl alcohol for fifteen minutes. The material waswashed with diethyl ether to give the title compound (0.715 g, 40%) asan off-white powder (m.p. 245-248° C.). ¹H NMR (300 MHz, DMSO-d₆): δ1.74 (d, 3H, J=6.9 Hz); 2.31 (s, 3H, CH ₃SO₃H); 6.24 (q, 1H, J=6.9 Hz);7.45 (dd, 1H, J=1.8, 8.7 Hz); 7.86 (m, 2H); 8.05 (d, 1H, J=1.8 Hz); 8.14(d, 1H, J=8.7 Hz); 8.82 (m, 2H); 12.03 (s, 1H); 12.71 (s, 1H). Calc'd.for C₁₈H₁₃ClN₄ _(O) ₃.CH₃SO₃H.0.8 H₂O: C, 47.61; H, 3.91; N, 11.68.Found: C, 47.84; H, 3.79; N, 11.54.

(+/−)-7-Chloro-4-hydroxy-2-(1-pyrid-4-yl-N-oxide)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione.

To a stirred suspension of(+/−)-7-chloro-4-hydroxy-2-(1-pyrid-4-yl)ethyl-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dionemethanesulfonate (0.400 g, 0.8 mmol) in methyl alcohol (1.5 mL) wasadded choline hydroxide (0.49 mL, 45% in MeOH, 1.73 mmol) and thesolution stirred until all had dissolved. To this was added3-chloroperoxybenzoic acid (0.316 g, 57-86% purity, 1.04 mmol-1.57 mmol)and the solution stirred 3 hours. The solid was then filtered, washedwith methyl alcohol and dried in vacuo (200 mtorr) to give the titlecompound (0.176 g, 57%) as a yellow solid. ¹H NMR (300 MHz, DMSO-d₆): δ1.65 (d, 3H, J=6.9 Hz) 6.14 (d, 1H, J=6.9 Hz); 7.30 (d, 2H, J=6.6 Hz);7.42 (d, 1H, J=8.7 Hz); 8.03 (s, 1H); 8.14 (m, 3H); 11.92 (br s, 1H);12.61 (br s, 1H). MS (+CI) m/z 385/387.

Tests for Biological Function:

Test A: Inhibition of Binding of [³H]-MDL105,519:

Binding of compounds to the NMDA receptor glycine site may be assessedby measuring the ability of test compounds to inhibit the binding oftritiated MDL105,519 to brain membranes bearing the receptor.

Rat Brain Membranes: The rat brain membranes used in the experimentswere obtained from Analytical Biological Services Inc., and wereprepared substantially in accordance with the method of B. M. Baron etal., J. Pharmacol. Exp. Ther. 250, 162 (1989). Briefly, fresh braintissue including cerebral cortex and hippocampus from male SpragueDawley rats was homogenized in 0.32 M sucrose and centrifuged at lowspeed to separate cellular membranes from other cellular components. Themembranes were then washed 3 times using deionized water, followed bytreatment with 0.04% Triton X-100. Finally, membranes were washed sixtimes in 50 mM Tris citrate buffer, pH 7.4, and frozen at −80° C. untiluse.

[³H]MDL105,519 (72 Ci/mmol) was purchased from Amersham. Cold MDL105,519was purchased from Sigma/RBI. Binding assays were performedsubstantially in accordance with the protocol of B. M. Baron et al., J.Pharmacol. Exp. Ther. 279, 62 (1996), as follows. On the day of theexperiment, brain membranes were thawed at room temperature andsuspended in 50 mM tris acetate buffer, pH 7.4 (“TAB”). Seventy-fivemicro grams per milliliter protein (by using the BioRad dye) were usedfor competition binding. The experiments were carried out using 96-wellplates. Membranes were incubated with 20 μL of compounds of variousconcentrations and 1.2 nM [³H]ML105,519 for 30 minutes at roomtemperature in a total volume of 250 μL. Non specific binding wasdetermined by using 100 μM of unlabeled MDL105,519. The unlabeledMDL105,519 and compounds were dissolved as 12.5 mM stock solutions inDMSO. Final DMSO concentration in each well was kept below 1%, whichconcentration was found not to alter the binding results. Afterincubation, unbound [³H]ML105,519 was removed by filtration onto GF/BUnifilter plates using a Packard harvester. Filters were washed fourtimes with ice cold TAB (total of 1.2 mL buffer). The plates were driedovernight at room temperature and bound radioactivity was measured on aPackard TopCount after the addition of 45 μL per well of the MICROSCINTO.

Human Brain Membranes: Human brain membranes were obtained fromAnalytical Biological Services Inc., and assays were performed asdescribed for rat membranes.

Data analysis: Data was analyzed using a Microsoft Excel spreadsheet andGraphPad Prizm software and potency of compounds is expressed as the Ki(nM).

Test B: Formalin Test:

The Formalin test is an assay that assesses the capacity of a compoundto inhibit formalin-induced nociceptive behaviors in rats (D. Dubuisson,et al., Pain 4, 161-174 (1977); H. Wheeler-Aceto et al.,Psyclopharmacology 104, 35-44 (1991); T. J. Coderre, et al., Pain 54,43-50 (1993)). In the test, two distinctive phases of formalin-inducedbehaviors are observed. A first phase response, caused by acutenociception to the noxious chemical (formalin) injected into the paw,occurs between zero and five minutes. A quiescent period of 5 to 15 minpost injection follows. After the quiescent period a second phaseresponse, caused by sensitization of the central neurons in the dorsalhorn, occurs after 15 minutes and lasts up to 60 minutes. Sensitizationof the central neurons in the spine augments a noxious afferent inputand causes a stronger pain barrage to be transmitted to the brain.Therefore, inhibition of the second phase response indicates a centralmechanism of drug action.

The procedure for the formalin test may be performed as follows: malerats are placed in a plexiglass chamber and observed for 30-45 min. toobserve their baseline activity. Animals would either be pretreated withvehicle or with different doses of a test compound and are dosed withvehicle or test compound three hours prior to injection of 0.05 mL ofsterile 1% formalin under the dorsal skin of a hind paw. The number ofpaw flinches (responses) during the first phase (0-5 min.) and thesecond phase (20-35 min.) are scored and recorded. Flinch response canbe compared with the mean score of a saline control group and calculatedas percentage inhibition. The ED₅₀ is the dose of compound whichproduced 50% inhibition of nociceptive response in the first or secondphase response.

% inhibition of nociceptive response can be calculated as:100×(number of responses in vehicle group−number of responses incompound group)/(number of responses in compound group)

Student's t-test can be used for statistical analysis to determine thesignificance of compound effects.

Test C: Neuropathic Pain Model (Chronic Constriction Injury):

The anti-hyperalgesic properties of a compound may be tested with theChronic Constriction Injury (“CCI”) model. The test is a model forneuropathic pain associated with nerve injuries that can arise directlyfrom trauma and compression, or indirectly from a wide range of diseasessuch as infection, cancer, metabolic conditions, toxins, nutritionaldeficiencies, immunological dysfunction, and musculoskeletal changes. Inthe model a unilateral peripheral hyperalgesia is produced in rats bynerve ligation (G. J. Bennett, et al., Pain 33, 87-107 (1988)).

Procedurally, Sprague-Dawley rats (250-350 g) are anesthetized withsodium pentobarbital and the common sciatic nerve exposed at the levelof the mid thigh by blunt dissection through the biceps femoris. Asection of nerve (about 7 mm), proximal to the sciatic trifucation, isfreed of tissue and ligated at four positions with chromic gut suture,with the suture tied with about 1 mm spacing between ligatures. Theincision is closed in layers and the animals allowed to recuperate.Thermal hyperalgesia is measured using a paw-withdrawal test (K.Hargreaves, et al., Pain 32, 77-88 (1988)). To perform the test, animalsare habituated on an elevated glass floor and a radiant heat sourceaimed at the mid-plantar hindpaw (sciatic nerve territory) through theglass floor with a 20 second cut-off to prevent injury to the skin. Thelatencies for the withdrawal reflex in both hind paws are recorded.

In this test, paws with ligated nerves show shorter paw withdrawallatencies compared to the unoperated or sham operated paws. Responses totest compounds are evaluated at different times after oraladministration to determine the onset and duration of compound effect.When performing the test, groups of CCI rats would receive eithervehicle or the test compound orally three times daily for 5 days. Pawwithdrawal latencies can be measured each day 10 min. before and two orthree hr. after the first daily dose. Compound efficacy is calculated asmean percentage decrease of hyperalgesia compared to a vehicle-treatedgroup. Compound potencies may be expressed as the minimum effective dose(MED) in mg/Kg/day that yields a % decrease in hyperalgesia that isstatistically significant, where the % anti-hyperalgesic effect may becalculated as follows:

 (Mean of vehicle group−Mean of compound group)×100/(Mean of vehiclegroup)

Data analysis can be performed by the multiple means comparison(Dunnett's test).

Table 1 shows the results from Test A for certain compounds of theinvention.

TABLE 1 Test A Ki (nM) Ex. 1  996 Ex. 2  751 Ex. 3  >10 μM Ex. 4  3690Ex. 5  907 Ex. 7  1780 Ex. 8  228 Ex. 10 2570 Ex. 11 2420

1. A compound according to structural diagram I;

wherein: R¹ is halo; A is CH; E is selected from phenyl andC₃₋₇cycloalkyl; D is selected from pyridyl and N-oxide of pyridyl, ortautomers or pharmaceutically-acceptable salts thereof.
 2. A compoundaccording to claim 1, wherein: E is selected from phenyl andC₃₋₅cycloalkyl.
 3. A compound of claim 1, according to structuraldiagram II:


4. A compound according to claim 3, wherein: E is selected from phenyland C₃₋₅cycloalkyl.
 5. A compound according to claim 1, selected from:7-chloro-4-hydroxy-2-(1-(1-cyclopropyl-1-pyrid-2-ylmethyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione;7-chloro-4-hydroxy-2-(1-cyclopropyl-1-pyrid-4-ylmethyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione;7-chloro-4-hydroxy-2-(1-cyclopentyl-1-pyrid-2-ylmethyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione,and7-chloro-4-hydroxy-2-(1-phenyl-1-pyrid-4-ylmethyl)-1,2,5,10-tetrahydropyridazino[4,5-b]quinoline-1,10-dione.6. A method for treating a subject suffering from pain comprisingadministering a pain-ameliorating effective amount of a compoundaccording to structural diagram I,

wherein: R¹ is halo; A is CH; E is selected from phenyl andC₃₋₇cycloalkyl; D is selected from pyridyl and N-oxide of pyridyl, ortautomers or pharmaceutically-acceptable salts thereof.
 7. A methodaccording to claim 6, wherein in a compound according to structuraldiagram I: E is selected from phenyl and C₃₋₅cycloalkyl.
 8. Apharmaceutical composition comprising a pain-ameliorating effectiveamount of a compound according to structural diagram I:

wherein: R¹ is halo; A is CH; E is selected from phenyl andC₃₋₇cycloalkyl; D is selected from pyridyl and N-oxide of pyridyl, ortautomers or pharmaceutically-acceptable salts thereof, together with apharmaceutically-acceptable excipient or diluent.